Comprehensive Resistome Analysis

KPC-31 Carbapenemase-Producing Enterobacterales

Author

Russell Lewis

Published

December 27, 2025

Executive Summary

This report presents a comprehensive analysis of 42 bacterial isolates from the KPC-31 resistome project. The dataset includes detailed characterization of carbapenemase-producing Klebsiella pneumoniae and Escherichia coli isolates, with emphasis on:

  • Species and molecular typing (sequence types, capsular types)
  • Antimicrobial resistance phenotypes (MIC values for 26 antibiotics)
  • Resistance genotypes (36 resistance genes identified)
  • Plasmid incompatibility groups (15 Inc groups detected)
  • Horizontal gene transfer patterns

Key Findings

ImportantCritical Discovery: Universal blaKPC-31 Carriage

ALL 42 isolates (100%) carry the blaKPC-31 carbapenemase gene, indicating this is a KPC-31-specific collection or outbreak. This variant is responsible for the observed carbapenem resistance and variable ceftazidime-avibactam (CAZ-AVI) resistance across isolates.

  • Dominant species: Klebsiella pneumoniae (78.6%, n=33)
  • Most prevalent sequence type: ST307 (50% of typed isolates, n=16)
  • High-risk clones identified: ST307, ST512, ST101, ST131, ST147
  • CAZ-AVI resistance: 35.7% show high-level resistance (MIC >16 mg/L)
  • Plasmid diversity: 15 incompatibility groups, with IncF plasmids spreading across multiple clones

1 Dataset Overview

1.1 Data Structure

Dataset Dimensions:
==================
• Total isolates: 42
• Total variables: 155
• Antibiotics tested (MIC): ~26
• Resistance genes analyzed: 36
• Plasmid Inc groups detected: 15

1.2 Data Completeness

Table 1: Data completeness for key metadata variables
Variable Total Complete Missing % Complete
ID 42 42 0 100.0
Species 42 42 0 100.0
Clinical Source 42 32 10 76.2
Sequence Type (ST) 42 32 10 76.2
Capsular Type (KL) 42 10 32 23.8
Wzi Type 42 9 33 21.4

2 Bacterial Characterization

2.1 Species Distribution

Figure 1: Distribution of bacterial species in the collection
NoteSpecies Composition
  • Klebsiella pneumoniae: 33/42 isolates (78.6%)
  • Escherichia coli variants: 9/42 isolates (21.4%)
    • E. coli TOP10: 5 isolates (lab strain)
    • E. coli DH5-α: 2 isolates (lab strain)
    • E. coli: 2 isolates (clinical)

2.2 Sequence Types (MLST)

Figure 2: Distribution of sequence types among typed isolates
Table 2: Summary of sequence types and their clinical significance
ST n Percentage Species Clinical.Significance
ST307 16 50.0 K. pneumoniae Global high-risk clone
ST101 7 21.9 K. pneumoniae MDR clone
ST512 5 15.6 K. pneumoniae Emerging high-risk clone
ST131 2 6.3 E. coli Global E. coli pandemic clone
ST147 2 6.3 K. pneumoniae MDR K. pneumoniae clone

2.3 Clinical Sources

Figure 3: Distribution of clinical specimen types

2.4 Capsular Types (KL)

ImportantPerfect KL-ST Association

Each capsular type is exclusively associated with one sequence type, suggesting stable clonal lineages:

  • KL107 ↔︎ ST512 (n=4)
  • KL102 ↔︎ ST307 (n=2)
  • KL17 ↔︎ ST101 (n=2)
  • KL64 ↔︎ ST147 (n=2)

Only 23.8% (10/42) of isolates have capsular typing data, but the perfect association is striking.


3 Antimicrobial Resistance Phenotypes

3.1 Ceftazidime-Avibactam (CAZ-AVI) Resistance

Figure 4: Distribution of CAZ-AVI MIC values

3.2 CAZ-AVI Resistance by Sequence Type

Figure 5: CAZ-AVI resistance stratified by sequence type
WarningST307 Heterogeneous Resistance

ST307 shows a 50/50 split between intermediate and resistant isolates, suggesting:

  • Evolving resistance mechanisms within this clone
  • Potential for further resistance development
  • Need for careful monitoring

3.3 Multi-Antibiotic Resistance Heatmap

Figure 6: Median MIC values across antibiotics by sequence type

4 Resistance Genotypes

4.1 blaKPC-31: The Universal Carbapenemase

Figure 7: blaKPC-31 prevalence across species and sequence types
ImportantUniversal KPC-31 Carriage

100% of isolates (42/42) carry blaKPC-31, including:

  • All K. pneumoniae isolates
  • All E. coli isolates (including lab strains)
  • All sequence types
  • Both clinical and environmental isolates

This indicates a KPC-31-specific collection or outbreak.

4.2 Resistance Gene Profiles by Clone

Figure 8: Resistance gene profiles for isolates with capsular typing
Table 3: Summary of resistance genes by KL/ST combination
Clone Total.Genes Beta.lactamases Key.Features
KL64 (ST147) 15 KPC-31, CTX-M-15, TEM-1B, SHV-11, OXA-1 Most genes; fosfomycin resistance
KL102 (ST307) 13 KPC-31, CTX-M-15, TEM-1B, SHV-28, OXA-1 qnrB1 quinolone resistance
KL107 (ST512) 12 KPC-31, TEM-1A, SHV-158 IncX3 plasmid-associated
KL17 (ST101) 5 KPC-31, SHV-1 armA (pan-AMG resistance)
NoteNotable Findings
  • KL64 (ST147): Highest gene count (15), includes fosA (fosfomycin resistance)
  • KL17 (ST101): Lowest gene count (5), but includes armA conferring pan-aminoglycoside resistance
  • KL102 (ST307): Carries qnrB1 providing quinolone resistance
  • KL107 (ST512): Associated with IncX3 plasmid

5 Capsular Type-Resistance Associations

5.1 KL-ST Perfect Association

Figure 9: Perfect 1:1 association between capsular types and sequence types

5.2 CAZ-AVI Resistance by Capsular Type

Figure 10: CAZ-AVI resistance profiles differ dramatically by capsular type
Table 4: CAZ-AVI resistance profiles by capsular type
KL.Type ST n Median.MIC Resistance.Status Clinical.Concern
KL64 ST147 2 256 100% Resistant Very High
KL17 ST101 2 130 100% Resistant Very High
KL107 ST512 4 48 100% Resistant High
KL102 ST307 2 12 100% Intermediate Moderate

6 Plasmid Incompatibility Groups

6.1 Inc Group Distribution

Figure 11: Prevalence of major incompatibility groups

6.2 Inc Groups by Clone

Figure 12: Incompatibility group distribution by KL/ST combination
ImportantClone-Specific Plasmid Profiles
  • KL107 (ST512): IncX3 (100%) - fixed, likely carries blaKPC-31
  • KL17 (ST101): IncR (50%) - may carry armA gene
  • KL64 (ST147): IncF family (50%) - high HGT potential
  • KL102 (ST307): NO Inc groups detected - chromosomal integration?

6.3 Horizontal Gene Transfer Evidence

Figure 13: Plasmid diversity indicates horizontal gene transfer potential
Table 5: Evidence for horizontal gene transfer by Inc group
Inc.Group Distribution HGT.Evidence Significance
IncFIB(K) / IncFII(K) 4 different STs ★★★★★ High Active dissemination within K. pneumoniae
IncX3 ST512 only ★☆☆☆☆ None Clone-specific, stable, not spreading
IncR Primarily ST101 ★★☆☆☆ Limited May carry armA, limited to one clone
IncN Rare, isolated ★★★☆☆ Moderate Recent acquisition, broad host range potential

7 Integrated Analysis

7.1 Relationships Between Plasmids, Genes, and Resistance

Figure 14: Integrated view of plasmid complexity, gene burden, and resistance levels
NoteKey Paradoxes
  1. KL102 (ST307): Zero Inc groups but 13 resistance genes → Chromosomal integration?
  2. KL17 (ST101): High plasmid complexity (2.3) but only 5 genes → Quality over quantity (armA effect)
  3. KL64 (ST147): Most genes (15) and highest MIC (256) → Synergistic effects

7.2 Summary Table: Clone Characteristics

Table 6: Comprehensive summary of clone characteristics
Clone n CAZ.AVI Resistance.Genes Key.Genes Dominant.Plasmid HGT.Risk Clinical.Priority
KL107 (ST512) 4 Resistant (48) 12 blaKPC-31, blaTEM-1A IncX3 (100%) Low High
KL17 (ST101) 2 Resistant (130) 5 blaKPC-31, armA IncR (50%) Low Very High
KL64 (ST147) 2 Resistant (256) 15 blaKPC-31, CTX-M-15, fosA IncF (50%) High Very High
KL102 (ST307) 2 Intermediate (12) 13 blaKPC-31, CTX-M-15, qnrB1 None detected Unknown Monitor

8 Clinical Implications

8.1 Treatment Challenges

WarningExtremely Limited Treatment Options

All isolates carry blaKPC-31, resulting in:

  • Carbapenems: Compromised by carbapenemase
  • ⚠️ CAZ-AVI: Variable resistance (35.7% high-level)
  • ⚠️ Aminoglycosides: Compromised (especially ST101 with armA)
  • ⚠️ Quinolones: Resistance common (gyrA, ParC, qnrB1)

8.1.1 Potential Treatment Strategies

Table 7: Potential treatment options by clone type
Clone First.Line Second.Line Last.Resort Notes
KL107 (ST512) Cefiderocol Carbapenem + AVI Colistin IncX3-stable resistance
KL17 (ST101) Cefiderocol Plazomicin (not armA) Colistin armA limits aminoglycosides
KL64 (ST147) Combination therapy Carbapenem + fosfomycin Colistin High gene burden, test all options
KL102 (ST307) Carbapenems CAZ-AVI Colistin Best prognosis of 4 clones

8.2 Infection Control Priorities

8.2.1 High-Risk Clones for Isolation

  1. ST512 (KL107): IncX3 plasmid stable but high resistance
  2. ST101 (KL17): armA-mediated pan-aminoglycoside resistance
  3. ST147 (KL64): IncF plasmids with high HGT potential

8.2.2 Surveillance Recommendations

Priority Target Rationale
Critical IncF plasmid spread Spreading across 4 STs, HGT evidence
Critical blaKPC-31 variants Monitor for evolution (KPC-32, KPC-33, etc.)
High ST307 mechanism No Inc groups detected - chromosomal integration?
High Capsular typing Strong predictor of resistance phenotype
Moderate CAZ-AVI MICs Early detection of resistance evolution

9 Conclusions

9.1 Key Findings Summary

  1. Universal blaKPC-31 carriage (100%, 42/42) defines this collection
  2. ST307 is dominant (50% of typed isolates) but shows heterogeneous resistance
  3. Perfect KL-ST associations indicate stable clonal lineages
  4. IncF plasmids are disseminating across multiple K. pneumoniae clones
  5. Clone-specific resistance mechanisms:
    • ST512: IncX3-based (stable, not spreading)
    • ST101: IncR-based (armA-mediated pan-AMG resistance)
    • ST147: IncF-based (high HGT risk)
    • ST307: Unknown mechanism (requires investigation)

9.2 Critical Questions for Further Investigation

TipRecommended Next Steps
  1. Whole plasmid sequencing: Map complete genetic context of resistance genes
  2. ST307 mechanism: Investigate chromosomal vs cryptic plasmid integration
  3. IncX3 plasmid characterization: Complete sequence of ST512 blaKPC-31-bearing plasmid
  4. Temporal analysis: If collection dates available, track resistance evolution
  5. Epidemiological links: Investigate patient/ward connections for transmission chains
  6. Functional studies: Test plasmid transferability and fitness costs

9.3 Public Health Implications

ImportantOutbreak Potential

This collection represents either:

  • A KPC-31-specific outbreak with multiple introductions
  • A surveillance collection of endemic KPC-31 strains
  • Clonal expansion of resistant lineages with horizontal gene transfer

The presence of high-risk global clones (ST307, ST512, ST101, ST131, ST147) combined with active plasmid dissemination (IncF family) represents a significant public health concern requiring immediate infection control measures.


10 Methods

10.1 Data Source

  • Dataset: Progetto 1 (KPC-31) versione 2.xlsx
  • 42 bacterial isolates
  • 155 variables including:
    • Bacterial identification (species, MLST, capsular typing)
    • Antimicrobial susceptibility testing (MIC for ~26 antibiotics)
    • Resistance gene detection (36 genes)
    • Plasmid replicon typing (15 incompatibility groups)

10.2 Analytical Approach

All analyses were performed using R version 4.5.1 with the following packages:

  • tidyverse for data manipulation and visualization
  • readxl for Excel file import
  • ggplot2 for graphics
  • knitr and kableExtra for tables

10.2.1 Resistance Categorization

  • CAZ-AVI Susceptible: MIC ≤8 mg/L
  • CAZ-AVI Intermediate: MIC 9-16 mg/L
  • CAZ-AVI Resistant: MIC >16 mg/L

10.2.2 Statistical Notes

  • Percentages calculated on non-missing values
  • Median MIC used for heatmaps to reduce outlier effects
  • Log2 transformation applied for MIC visualization

11 Appendix

11.1 Complete Resistance Gene List

Table 8: Complete list of resistance genes detected in the dataset
Category Gene Mechanism
Beta-lactamases
Beta-lactamases BlaKPC-31 Carbapenemase (KPC)
Beta-lactamases BlaCTXM-15 ESBL (CTX-M)
Beta-lactamases blaTEM-1A Beta-lactamase (TEM)
Beta-lactamases blaTEM-1B Beta-lactamase (TEM)
Beta-lactamases blaSHV-1 Beta-lactamase (SHV)
Beta-lactamases blaSHV-11 Beta-lactamase (SHV)
Beta-lactamases blaSHV-28 Beta-lactamase (SHV)
Beta-lactamases BlaSHV-106 Beta-lactamase (SHV)
Beta-lactamases BlaSHV-158 Beta-lactamase (SHV)
Beta-lactamases blaOXA-1 Beta-lactamase (OXA)
Beta-lactamases bla OXA-9 Beta-lactamase (OXA)
Aminoglycosides
Aminoglycosides aadA Aminoglycoside acetyltransferase
Aminoglycosides aaaA2 Aminoglycoside resistance
Aminoglycosides aac(6′)-Ib' Aminoglycoside acetyltransferase
Aminoglycosides aac(6´)-Ib-cr AMG + FQ resistance
Aminoglycosides ant(2′')-Ia Aminoglycoside nucleotidyltransferase
Aminoglycosides aac(3)-IIa Aminoglycoside acetyltransferase
Aminoglycosides armA 16S rRNA methylase (pan-AMG)
Aminoglycosides aph(3')-Ia Aminoglycoside phosphotransferase
Quinolones
Quinolones ParC DNA gyrase mutation
Quinolones gyrA DNA gyrase mutation
Quinolones qnrB1 Quinolone resistance
Other Mechanisms
Other Sul1 Sulfonamide resistance
Other Sul2 Sulfonamide resistance
Other fosA Fosfomycin resistance

11.2 Abbreviations

  • AMG: Aminoglycoside
  • CAZ-AVI: Ceftazidime-avibactam
  • ESBL: Extended-spectrum beta-lactamase
  • HGT: Horizontal gene transfer
  • Inc: Incompatibility
  • KL: Capsular type (K locus)
  • MDR: Multi-drug resistant
  • MIC: Minimum inhibitory concentration
  • MLST: Multi-locus sequence typing
  • ST: Sequence type

11.3 Session Information

Code
sessionInfo()
R version 4.5.1 (2025-06-13)
Platform: aarch64-apple-darwin20
Running under: macOS Tahoe 26.3

Matrix products: default
BLAS:   /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRblas.0.dylib 
LAPACK: /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRlapack.dylib;  LAPACK version 3.12.1

locale:
[1] C.UTF-8/C.UTF-8/C.UTF-8/C/C.UTF-8/C.UTF-8

time zone: Europe/Rome
tzcode source: internal

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
 [1] kableExtra_1.4.0 knitr_1.50       readxl_1.4.5     lubridate_1.9.4 
 [5] forcats_1.0.1    stringr_1.6.0    dplyr_1.1.4      purrr_1.2.0     
 [9] readr_2.1.6      tidyr_1.3.1      tibble_3.3.0     ggplot2_4.0.1   
[13] tidyverse_2.0.0 

loaded via a namespace (and not attached):
 [1] gtable_0.3.6       jsonlite_2.0.0     compiler_4.5.1     tidyselect_1.2.1  
 [5] xml2_1.5.1         textshaping_1.0.4  systemfonts_1.3.1  scales_1.4.0      
 [9] yaml_2.3.12        fastmap_1.2.0      R6_2.6.1           labeling_0.4.3    
[13] generics_0.1.4     htmlwidgets_1.6.4  svglite_2.2.2      pillar_1.11.1     
[17] RColorBrewer_1.1-3 tzdb_0.5.0         rlang_1.1.6        stringi_1.8.7     
[21] xfun_0.55          S7_0.2.1           viridisLite_0.4.2  timechange_0.3.0  
[25] cli_3.6.5          withr_3.0.2        magrittr_2.0.4     digest_0.6.39     
[29] grid_4.5.1         rstudioapi_0.17.1  hms_1.1.4          lifecycle_1.0.4   
[33] vctrs_0.6.5        evaluate_1.0.5     glue_1.8.0         cellranger_1.1.0  
[37] farver_2.1.2       rmarkdown_2.30     tools_4.5.1        pkgconfig_2.0.3   
[41] htmltools_0.5.9   

Report generated on: December 28, 2025

Analysis platform: Positron 2025.12.2 with R 4.5.1